1a.Objectives (from AD-416)
Develop techniques and strategies that utilize molecular gene transfer methods to create transgenic strains of Diptera, Lepidoptera, and Coleoptera that will facilitate genetic-sexing or have novel autocidal properties for use in IPM programs. Identify strains of Lepidoptera pests, describe differences in behaviors, and isolate the genetic factors controlling these differences in order to understand how species adapt to new ecological niches and to better target biologically-based control strategies in area-wide IPM programs. Describe acoustic and other signals and cues produced by pest arthropods, including Mediterranean fruit fly, Diaprepes root weevil and other cryptic/hidden insects, and develop detection technologies and attractive devices that can be used in IPM projects to target, monitor and control pests. Develop strategies for effective use of parasitoids and predators in IPM of Lepidoptera and tephritid fruit fly pests, such as Mediterranean fruit fly, potentially invasive species of Anastrepha fruit flies and fall armyworm, through behavioral and ecological studies of their feeding, mating, dispersal and oviposition.Identify, isolate, and characterize biochemical factors from prey, or from cell lines derived from prey, that stimulate development, determine their physiological and molecular effects, and use these factors to improve artificial diets for mass-reared predators. .2)Measure the effects of optimal and suboptimal diets, including essential and alternative prey and artificial diets, on reproductions and predatory efficiency in laboratory and field-cage studies. Identify, isolate, and characterize biochemical factors from prey, or from cell lines derived from prey, that stimulate development, determine their physiological and molecular effects, and use these factors to improve artificial diets for mass-reared predators. Measure the effects of optimal and suboptimal diets, including essential and alternative prey and artificial diets, on reproductions and predatory efficiency in laboratory and field-cage studies.

1b.Approach (from AD-416)
Emphasis will be placed on.1)Using recombinant DNA constructs inserted into the piggyBac transformation vector to genetically transform strains of the fruit flies and moth ;.2)Developing a means of generating somatic transformations to test the phenotypes and efficiencies of foreign gene constructs that lead to genetic control of pest insects;.3)Assessing the potential for vector re-mobilization in released transgenic strains and developing new vectors that allow increased stability and targeted integration for greater efficacy and ecological safety;.4)Defining the seasonal distributions of genetically distinct subpopulations of fall armyworm in order to investigate strain-specific behaviors related to plant host usage, migration, and mating;.5)Identifying hidden/cryptic pests through acoustic and microwave radar technology and precisely targeting control measures to limited areas where they will be most effective;.6)Developing economical all-female strains of fruit fly parasitoids for mass-rearing and augmentative release through sex ratio distorting microbial endosymbionts; and.7)Determining how fruit fly and moth parasitoids differ in their abilities to locate hosts at varying densities, and display different propensities to disperse from areas with relatively low host-encounter rates.

Research is needed to enhance biological control of insects and mites through improved methods for rearing and deploying arthropod predators. The impact of predators can be increased through discovery of essential nutritional factors from prey or hosts that increase reproductive efficiency. Following bioassay-guided isolation and characterization, these factors will be employed as additives for artificial diets. Biochemical assays of contents of these factors in prey will also allow estimates of prey quality as food for predators. Biochemical and behavioral analyses of predators will produce estimates of fitness in insectary and field. Correlations of predator fitness with species and abundance of prey in field-cages will yield knowledge of the quality of prey as food and the quality of predators introduced into cages after rearing on selected diets in the laboratory. Predatory efficiency of generalists such as Orius insidiosus, Podisus maculiventris, and Geocoris punctipes will be assessed through predation rates and selection of prey. Their reproductive potential will be assessed as egg load. General fitness of the predators will be measured through quantitative and qualitative analyses of lipids and proteins. Results will yield commercial additives for artificial diets for predators, biochemical markers for assessment of insect quality, and tools for improved management of existing and introduced populations of predators and pest populations in the field and greenhouse.

3.Progress Report
Progress was made in the all the major goals of this project: identification of pest populations and their dispersal, discovery of new biological control agents, their mass-rearing and release. In particular, research advanced the creation of genetic modifications that can induce fruit fly male sterility for Sterile Insect Technique (SIT) programs, but which do not require debilitating irradiation of released males. Technology for inducing new genetic combinations through viruses was transferred to a number of other laboratories, and a new means of adding genes to the body of an insect has been developed. In collaboration with APHIS, a genetic marker that identifies a fruit fly as mass-reared and sterile was found to be stable in a number of trap types. Such experiments will lead to the effective field release of genetically modified control agents. The migrations of the fall army worm were described in finer detail and target areas for their area-wide management have been identified. Work continues on applying biological control to these “hot-spots”. Migration of fall army worms from overwintering sites is being used to model the effects of climate change on the distributions of pests. Chemical attractants are not useful in some situations where pests are hidden and immobile, such as when they infest, grain or wood, but their presence can still be discovered through the sounds they make as they feed. Acoustic pest identification techniques continue to be applied domestically and overseas in Saudia Arabia and Aruba/Curacao. Fruit fly parasitoid host-based oviposition stimulants were identified which could improve production in mass-rearing facilities. Conservation biological control is enhanced by providing food and shelter for natural enemies and continuing surveys have demonstrated that certain plants provide food and alternative hosts for both insect predators and parasitoids.

This project has termed and been replaced by project #6615-22000-024-00D.

4.Accomplishments
1.
Migration of Bt-insecticide resistant moths: Fall armyworm, have extensive northward migrations that begin with subtropical overwintering populations. The first example of field-evolved resistance to a Bacillus thuringiensis (Bt) insecticide recently occurred in Puerto Rican fall armyworm and the rapid spread of this resistance to the rest of the western hemisphere is plausible. Unfortunately, relatively little was known about the population movements of fall armyworm in the Caribbean. A novel method using mitochondrial genes was used to study migration of fall armyworm from Puerto Rico. The results indicate limited interactions between Puerto Rican fall armyworm and those from Brazil or Texas, but the potential for significant exchanges with populations in Florida. Monitoring Florida armyworms for resistance traits is recommended.

2.
RNAi for efficiency of gene silencing: The ability to silence, turn-off, genes critical for digestive activity in insects requires an efficient delivery of RNA “interference” molecules. A new means of making this delivery, a vector that is consumed by bacteria, can transform insect embryos or cultured insect cells. As a proof of principle, this vector was used to silence the production an otherwise fluorescing protein in insect cell cultures and now being combined with RNAi that will silence gut-specific genes in moths.

3.
Identification of beetle infestations in palm trees: Acoustic methods were developed to detect a newly invasive red palm weevil in Curacao and Aruba and will be used in monitoring and eradication programs. Signal processing methods were used that could distinguish the beetle from background noises and the sounds produced by many of the other nonpest insects commonly present in the trees. Use of this method will enable faster inspection of palm trees and reduce the numbers of trees that might be cut unnecessarily.

4.
New genes for lethality in fruit flies: Mass releases of sterile males are a widely used means to control pest fruit flies, but the radiation that sterilizes males often damages their sexual performance as well. Conditional-Lethality, where a released insect's offspring die when certain environmental conditions prevail, is a promising substitue for traditional sterility. Several new genes critical for conditional embryonic lethality have been discovered in the Mexican and Caribbean fruit flies and these could utimately improve the efficacy of control programs that protect US agriculture from not only fruit flies but also other potentially invasive pests.